Biology 101 2 25 99 1 st lecture for test two/thursday DNA Structure and Replication We have 23 paternal chromosomes and 23 maternal chromosomes from dad and mom. The chromosomes are chosen randomly on the chromatin chain by enzymes. Reason why we are unique. We have 3 billion nucleotides in DNA. The 5 Carbon sugar structure is part of the double helix structure. Know how to label all 5 sides and their base is made out of. C1 is the nitrogenous base, A, C, T, G or U on RNA. C2 is the sugar minus the oxygen atom. C3 is -OH and C4 is a phosphate. A connects to T; C connects to G The DNA nucleotide has three main groups, a nitrogenous base, a sugar, and a phosphate group. The nucleotides are joined by covalent bonds between the sugar of one nucleotide and the phosphate of the next. This results in a sugar phosphate backbone. DNA ligases ties the single DNA strand into one covalently bonded strip during replication. 1
DNA polymerases/replication DNA replication begins at specific sites on the double helix, called origins of replication. Then replication proceeds in both directions, creating replicating bubbles. The parental DNA strands open up as the daughter strands elongate on both sides of each bubble. Thousands of bubbles can be present at once. Eventually, all the bubbles merge, yielding two completed daughter DNA molecules. The DNA structure is universal; the sequence is unique to everyone else, except for twins. All have the same diameter of 2 nanometers. Each strand has a 3 1 (three-prime) end and a 5 1 end. The prime numbers refer to the carbon atoms of the nucleotide sugars. At one end of each DNA strand, the sugar s 3 prime carbon is attached to an OH group. At the other end, the sugar s 5 prime carbon has a phosphate group. The opposite orientation of the strands is important in DNA replication. The enzymes that link DNA nucleotides to a growing daughter strand, called DNA Polymerases, add nucleotides only to the 3 prime end, never to the 5 prime end. So 3 to 5, then the daughter strand can only grow in the 5 to 3 direction. 30 enzymes do this break up. She wants us to know two, DNA polymerases and DNA ligases. As one strand goes up the other goes down, in between are the DNA nucleotides connected by weak Hydrogen bonds. The strong covalent bonds are the backbone of the double helix (sugar covalently bonded to a phosphate, sugar phosphate sugar phosphate). This mirror image of the strands is called ANTI PARALELLISM. Then the two strands coil into the famous double helix. 2
Each strand is made up of an old and new strand called the Semi Constructive Model. There are minor and major grooves on the DNA ladder. The major grooves are fatter than the minor ones. Replication We re going to open up the DNA strand in the middle. During replication an enzyme arbitrarily chooses a sight on the strand the DNA opens up in the middle because of the weak H bonds. If the backbone were to break, some of the DNA would get lost. When we re finished we ll have two sets of chromosomes. 3
Biology 101 3 2 99 2 nd lecture for test two/tuesday Meiosis cell division in gametes, where the number of chromosomes is reduced by ½. Mitosis is for growth and repair, as in skin cells. It results in two daughter cells. It starts with one cell and gets two cells that are identical. Complete set is Meiosis involves only the sperm and eggs, the gametes and results in 4 cells. Each with 46 chromosomes (2n). n= 23 Haploid number 2n = complete chromosomes for humans. MITOSIS Chromosomes duplicate and then split in two as a cell divides. Before a eukaryotic cell begins to divide, it duplicates all of its chromosomes. The DNA molecule of each chromosome is copied. The result is that each chromosome now consists of two copies called Sister Chromatids. They are joined together at a specialized region called the centromere. The fuzzy appearance of the chromosome comes from the twists and folds of its chromatin fibers. When the cell divides, the sister chromatids of a duplicated chromosome separate from each other, Once separated from its sister, each chromatid is now called a chromosome and is identical to the chromosome we started with. In our own body, millions of cells must divide every second to maintain the total number of 60 trillion cells. Specialized cells such as nerve and muscle do not regenerate themselves. Eukaryotic cells undergo a CELL CYCLE, an orderly sequence that extends from the time a cell divides to form two daughter cells to the time those daughter cells divide again. 4
Most of the time, the cell cycle is spent in INTERPHASE. The cell s metabolic rate is very high chromosomes replicate during this time and serves as the basis for dividing interphase into three subphases, G1, S, G2 and M G1 is the period before DNA synthesis begins. G stands for gap. S is when DNA Synthesis occurs. At the end of this phase, the chromosomes are double each other. Each consisting of two sister chromatids. In a process called Mitosis, the nucleus and its contents, including the duplicated chromosomes, divide and are evenly distributed into two daughter nuclei. In a second process called cytokinesis, the cytoplasm divides in two. Together, these two cycles are called the mitotic phase, M of the cell cycle. MITOSIS: The Five Stages of Division 1. Interphase it is a period of growth when the cell synthesizes new molecules and organelles. The cell contains two microtubule organizing centers (MTOC s), which are clouds of cytoplasmic material that contain centrioles. 2. Prophase Within the nucleus, the chromatin fibers become more tightly coiled and folded, forming discreet chromosomes that can be seen easily. Each duplicated chromosome appears as 2 identical sister chromatids joined at the centromere. The mitotic spindle begins to form as microtubules grow out from the MTOC s which move to the poles. Then the nuclear envelope starts to disappear so the microtubles from both ends can reach into the chromosomes, now highly condensed. At the centromere region, each sister chromatid has a protein structure called the kinetochore. The spindle fibers attach on to this kinetochore. However some do not grab on. These are called the polar microtubules (PMTS). 3. Metaphase The mitotic spindle is fully formed, with its poles at opposite ends of the cell. The chromosomes convene on the metaphase plate, right in the center of the cell. The KMT attached to a particular chromatid all come from one pole of the spindle. And those attached to its sister chromatid come from the opposite pole. 4. Anaphase Begins when the two centomeres of each chromosome come apart, separating the sister chromatids. Once separated, each sister chromatid is now a fullfledged daughter chromosome. Anaphase is over when the equivalent and complete collections of chromosomes have reached the poles of the cell. 5. Telophase & Cytokinesis The end of cell division. Roughly the reverse of prophase. The nuclear envelope starts to reappear and the chromatin fiber of each chromosome uncoils itself. The mitotic spindle also disappears. Cytokinesis, the division of the cytoplasm, occurs along with telophase, with 2 daughter cells completely separating at the end of mitosis. 5
Review: On test two, this will be matching. Identify G1, working and growth. It stands for gap. S is DNA synthesis. DNA replication takes place before cell division. G2 is when the cell prepares to divide. It spans the time from completion of DNA synthesis to the onset of cell division. G2 is also a time of metabolic activity. Know these points. The plasma nuclear membrane disappears in late prophase. The centromere will be the region where the kinetochore lies. The kinetochore is the handle the MT will grab a hold on to. The kinetochore reinforces the DNA. There are two kinds of microtubules. The kinetochore microtubules which attach to the centromere and the polar microtubules are the ones that don t grab on to them. The polar MT makes the membrane become more oval because they are getting longer. Going into prophase, when the nuclear membrane is gone, the MT attaches on to the kinetochore. The chromosomes are going to wind up into little balls. We are shortening and condensing the DNA. This is why we can see the X structure so clearly. 6
Continue review: In metaphase, a short phase, some of the MT grab onto the kinetochore. Those that don t, the polar MT, will go to the poles and overlap each other. All the DYADS are lined up in the middle. In anaphase, the chromosomes come apart, separating the sister chromatids. The PMT get longer and longer and the KMT will get shorter and shorter as the pull the chromosomes to the poles. In telophase, the chromosomes unwind and the nuclear membrane reforms. The microtubules will fall into their tubulin form. In cytokinesis, ACTIN separates the cell into two. On test 2, explain the importance of 46 DYADS in metaphase in mitosis. It s so we have the correct 46 chromosomes in each cell. 7
Biology 101 3 4 99 3 rd lecture for test two/thursday MEIOSIS Meiosis is the reproduction of gametes, sperm and ova. The result is four haploid cells with 23 chromosomes each. An egg and sperm have 23 chromosomes each, so a zygote will have 46 chromosomes. Cells whose nuclei contain 2 homologous sets of chromosomes are diploid cells, and the total number of chromosomes is called the diploid number (2n). In humans the diploid number is 46. Under a microscope, we can see our chromosomes match up as 23 pairs. They are similar in length, centromere position and staining pattern. Metaphase chromosomes can be matched up and called homologous chromosomes. They are called homologous because they both carry GENES controlling the same inherited characteristics. The locus is a particular point for both chromosomes that have this gene for a trait. For example, eye color. We inherit one chromosome of each homologous pair from our mother and the other chromosome of the pair from our father. Our 23 pairs of homologous chromosomes are of two general types. 22 pairs consist of autosomes, found in both males and females. The other pair of chromosomes are the GAMETES, the sex cells. XX female. XY male. Each gamete has a single set of chromosomes, 22 autosomes plus a single sex chromosome, either X or Y. A cell with a single set is called a haploid cell, n=23. 8
During sex, a haploid sperm cell fuse with a haploid egg cell of the mother during fertilization. Results in a zygote, with a diploid number of 46. Mitotic cell division ensures that all somatic cells of the body receive copies of all the zygote s 46 chromosomes. Having haploid gametes keeps the chromosome number from doubling in each succeeding generation. 46, 92, 184 We don t have a human with 184 chromosomes. Haploid gametes are produced by a special sort of cell division called meiosis, which occurs only in reproductive organs. The cells divide into four cells either in the female ovaries or in the male s testes. So, whereas mitosis produces daughter cells with the same numbers of chromosomes as the parent cell, meiosis reduces the chromosome number by half. In meiosis, a cell undergoes 2 consecutive divisions, called meiosis I and meiosis II. Four daughter cells result from these divisions. MEIOSIS I Interphase I chromosomes duplicate here. At the end of interphase, each chromosome consists of 2 genetically identical sister chromatids. Prophase I most complex. Accounts for 90% of meiosis. In a process called SYNAPSIS, homologous chromosomes (each composed of two sister chromatids) pair up. They form a structure called a TETRAD. Each TETRAD has four chromatids. During synapses, chromatids of homologous chromosomes exchange segments, in a process called CROSSING OVER. VARIABILITY #I takes place here. Then the MTOC s (microtubule organizing centers) move away from each other and a spindle starts to form between them. The nuclear envelope disappears and the chromosomes tetrads, captured by spindle microtubules, are moved toward the center of the cell. The random alignment of maternal or paternal DYADS is called INDEPENDENT ASSORTMENT. This is to ensure everyone is different. A TETRAD IS TWO DYADS. Metaphase I The random attachment to the kinetochore results in the randomness of variability. Each chromosome is condensed and thick, with its sister chromatids attached at their centromeres by spindle microtubules. VARARIABLITY #2 takes place here. Anaphase I the chromosomes are then moved toward opposite ends of the cell. In contrast to mitosis, the sister chromatids making up each doubled chromosome remain attached at their centromeres. The dyads are still held together at their 9
kinetochore. Only the TETRADS, pairs of homologous chromosomes split up. Thus, you still see the 2 doubled chromosomes, the haploid #, moving toward each spindle pole. Telophase & cytokinesis In telophase I, the chromosomes arrive at the poles of the cell. When the chromosomes finish their journey, each pole of the cell has a haploid chromosome set. Usually cytokinesis occurs along with telophase I and the two haploid daughter cells are formed. No chromosomes duplication ocurrs between telophase I and the onset of meiosis II. MEIOSIS II After a pause in meiosis I, the chromosomes condense again and the nuclear envelope breaks down during prophase II. In any case, meiosis II is essentially the same as mitosis. Meiosis II separates the sister chromatids of the haploid number of chromosomes in the two starting cells, resulting in 4 daughter cells that have the haploid number of single daughter chromosomes. The important difference is that meiosis II starts with 2 haploid cells (23 chromosomes in each) that divide and produce 4 daughter cells with 23 chromosomes each. Lecture: We have tetrads in meisosis and dyads in mitosis They ll be a short answer question on meiosis on test 2 10
3 9 99 /4 th lecture for test two/tuesday DNA Transcription in the nucleus and Translation in the ribosomes. Transcription produces genetic messages in the form of RNA. Transcription, the transfer of genetic information from DNA to RNA, occurs in the cell nucleus. As with replication the two DNA strands unwind but only one-strand serves as a template for the newly forming RNA strand. The RNA nucleotides are linked by the transcription enzyme RNA Polymerase. The start transcribing signal is a nucleotide sequence called a promoter, which is located in the DNA. The first phase of transcription, called initiation, occurs when RNA polymerase attaches to the promoter DNA. INTIATION. During the 2 nd phase of transcription, the RNA elongates. Then in the 3 rd phase, termination, the RNA polymerase reaches a special sequence of bases in the DNA template called a terminator. This sequence signals the end of the gene on the DNA! So the RNA strand detaches from the gene AND the RNA polymerase to go outside the nucleus into the cytoplasm. ELONGATION AND TERMINATION AND THE COMPLETED RNA STRAND. There are 3 types of RNA Messenger RNA the ticker tape message that goes to the ribosome to be translated Transfer RNA the interpreter of the MRNA message that puts down the correct bases on the constructing protein strand. Ribosome RNA the stuff that RNA is made out of. RIBOSOMES build polypeptides A ribosome consists of 2 subunits, each made up of proteins and rrna. Its large subunit has binding site for trna and the small subunit binds for mrna. On the large subunit are the P and A sites. The P site is where holds the trna carrying the growing polypeptide chain, while the A site holds the trna carrying the next amino acid to be added to the chain. The A guy hands over the bases, (AUG etc.) over to the P site who actually puts the chain together by the trna. 11
An initiation codon marks the start of an mrna message. Translation can be divided into the same three phases as transcription: initiation, elongation and termination. An mrna molecule binds to a small ribosomal subunit. A special initiator trna locates and binds to the specific codon, called the start codon, where translation is to begin on the mrna molecule. A special sequence of nucleotides on a gene are not part of the message but helps the mrna bind to a ribosome. The role of the initiation process is to determine exactly where translation will begin, so that codons on the mrna are translated into the correct sequence of amino acids. ELONGATION ADDS AMINO ACIDS TO THE POLYPEPTIDE CHAIN Until a stop codon terminates translation. Once initiation is complete, amino acids are added one by one to the initial amino acid. Each addition occurs in a 3-step elongation process. Step one Codon recognition. The anticodon of an incoming trna molecule, carrying its amino acid, pairs with the mrna codon in the A site of the ribosome. The A site has the amino acids that are the opposite of the mrna chain. So if the mrna has AUG then the anticodon translation will be UAC respectively. Step two Then comes the PEPTIDE BOND formation. The polypeptide separates from the trna to which it was bound, the one in the P site, and attaches by a peptide bond to the amino acid carried by the trna in the A site. A first then P. Translocation the P site now leaves the ribosome and the A site trna, carrying the growing polypeptide, is translocated to the P site. A moves to P. Elongation continues until a stop codon reaches the ribosomes A site. Stop codons, UAA, UAG, UGA and ACT do not code for amino acids but instead signal translation to stop. The completed polypeptide is freed from the last trna and from the ribosome, which splits into its subunits. During and after translation, the polypeptide coils and folds into a tertiary and then later forming a quaternary structure. 12
MUTATION A change in the DNA sequence that affects the structure of the proteins. Almost all the nonsense amino acids, 70%, will be miscoded and usually new stops result in the codon. In cystic fibrosis, the condition can be traced back through the difference in a protein to one tiny change in a gene. In the hemoglobin molecule, the sickle cell child has a single different amino acid, a Val, instead of a Glu. This difference is caused by the change of a single nucleotide in the coding strand of DNA. We now know that the alternative alleles of many genes result from changes in single base pairs in DNA. Any change in the nucleotide sequence of DNA is called a mutation. It can involve large regions of a chromosome or just a single nucleotide. Mutations within a gene can be divided into two general categories: Base Substitutions and Base Deletions. A base substitution is the replacement of one nucleotide with another. If an mrna codon is GUA, instead of GAA, then CAT, mutant hemoglobin, results instead of CTT, normal hemoglobin. It may result in no change at all or an insignificant amount or life threatening. Mutations involving the insertion or deletion of one or more nucleotides in a gene often have disastrous effects. Because mrna is read as a series of nucleotide triplets during translation, adding or subtracting nucleotides may alter the reading frame (triplet grouping) of the genetic message. All the nucleotides that are downstream of the insertion or deletion will be regrouped into different codons. The result will most likely be a no0nfunctional polypeptide. 13
3 11 99 /5 th lecture for test two/thursday INHERITANCE Genetics What you see expressed. The genes that are used by the organism. In contrast to the Genotype All the genes of any organism that are passed down through offspring. Gregor Mendel discovered the fundamental principles of genetics breeding garden peas. The paternal parents are called the P Generation, and their HYBRID offspring are the F1 generation. Then when the F1 offspring mate, they produce the F2 generation. A monohybrid cross has one trait, while a dihybrid cross has 2 traits. Mendel s Hypothesis There are alternative forms of genes, the units that determine heritable traits. Ex. The gene for flower color exits in one form for purple and another for white. We now call alternative forms of genes ALLELES. For each inherited characteristic, an organism has two genes, one from each parent. These genes may both be the same allele or they may be different alleles. A sperm or egg carries only one allele for each inherited trait, because allele pairs separate from each other during the production of gametes. When they unite, each contributes its allele, thus restoring the paired condition in the offspring. When the two genes of a pair are different alleles, one is fully expressed and the other has no noticeable effect on the organism' appearance. These are called the DOMINANT ALLELE and the RECESSIVE ALLELE respectively. A true breeding organism, which has a pair of identical alleles for a characteristic, is said to be homozygous for that characteristic. Ex. AA or aa. While an organism with 2 different alleles for a characteristic, such as a pea plant with alleles P and p, is said to be heterozygous for that characteristic. There is a 3:1 ratio in the F2 generation, the Punnet Square shows the 4 possible combinations. The Punnet Square indicates the proportions of F2 plants predicted by Mendel s hypotheses. Because an organism s appearance does not always reveal its genetic composition, geneticists distinguish between an organism s expressed traits, PHENOTYPE, and its genetic make up, GENOTYPE. One parental trait disappears in the F1 generation of heterozygous, only to reappear in ¼ of the F2 offspring. The mechanism underlying this pattern is stated by Mendel s PRINCIPLE OF SEGREGATION Pairs of genes segregate during gamete formation, the fusion of gametes at fertilization pairs genes once again. 14
MENDEL S PRINCIPLE OF INDEPENDENT ASSORTMENT Each characteristic is independently inherited from the parents. The gametes RY and ry could produce four types of gametes, RY, ry, Ry and ry in equal quantities. The Punnet Square shows all the possible combinations of alleles that can result in the F2 generation from the union of four kinds of sperm with four kinds of eggs. The Punnet Square shows that each pair of alleles segregates independently during gamete formation. This behavior of genes is called Mendel s Principle of Independent Assortment. MANY GENES HAVE MORE THAN 2 ALLELES Many genes have multiple alleles. Although each individual carries, at most, two different alleles for a particular gene, in cases of multiple alleles, more than 2 possible alleles exist. The ABO blood group groups in humans are one example of multiple alleles. There are four phenotypes for this characteristic: A person s blood type may be O, A, B or AB. O is the universal donor, he produces no sugar. AB can take anybody s blood but he can only take AB blood. A single gene may affect many phenotypic characteristics. The impact of a single gene on more than one characteristic is called PLEITROPY. In many cases, one gene influences several characteristics. An example is sickle cell disease, where a single allele causes numerous health problems because it makes RBC produce abnormal hemoglobin molecules. These cells are destroyed by the body, lowering the RBC count causing anemia and general weakening of the body. It kills 100,000 people in the world annually. A single characteristic may be influenced by many genes Mendel studied genetic characteristics that could be classified on an either or basis, such as purple or white flower color in peas. However, many characteristics, such as human skin color and height, vary in population along a continuum. Many such features result from polygenic inheritance, the additive effect of two or more genes on a single phenotypic characteristic. This is the converse of pleitropy. 15
3 16 99 /6 th lecture for test two/tuesday HOW ONLY CERTAIN GENES ON DNA ARE EXPRESSED In both eukaryotes and prokaryotes, cell specialization depends on the selective expression of genes. The overall process by which genetic information flows from genes to proteins, from genotype to phenotype is called gene expression. E. COLI When we drink milk, we take in lactose. E. coli makes enzymes necessary to absorb the sugar and use it as an energy source. E. coli can make lactose utilization enzymes because it has genes that code for these enzymes. E. coli uses three enzymes to take up and start metabolizing lactose and the genes coding for these enzymes are regulated as a unit. Adjacent to the group of lactose enzyme genes are short sections of DNA that help control them. One stretch of nucleotides is a promoter, a site where the transcription enzyme, RNA polymerase, attaches and initiates transcription. Between the promoter and the enzyme genes, a DNA segment called an operator acts as a switch. The operator determines whether RNA polymerase can attach to the promoter and move along the genes. Such a cluster of genes with related functions, along with a promoter and an operator, is called an operon, operons exist only in prokaryotes. 16
3 18 99 /7 th lecture LAST LECTURE for Test two /Thursday RECOMBINANT DNA TECHNOLOGY In the mid-70 s, research on the E. Coli bacteria led to the development of recombinant DNA technology. This is a set of techniques for combining genes from different sources even different species and transferring it into cells. Scientists have already created genetically engineered bacteria that can mass-produce useful chemicals, including hormones and certain cancer drugs. 17